Healing of craniofacial defects is typically accomplished through the procedure of Distraction Osteogenesis. In the procedure, linear stresses are applied to osteoblasts and osteoblast precursors as the ends of the bone defect are pulled/distracted outward. This linear stress in known to have significant effects on the osteogenic capacity of the pre-osteoblast population - increasing their ability to differentiate and form new bone. In essence, linear forces may "pre-condition" progenitor cells and enhance their ability to differentiate into bone. In this grant application, the principle of distraction is applied to stem cell differentiation and their repair of bony defects. Specifically, this application attempts to determine if the application of linear forces can pre- condition adipose-derived stem cells (ASCs) and enhance their ability to heal bony defects. Therefore, this application describes the potential use of a novel in vitro distraction device that may impose similar mechanical stresses at a cellular level as those seen using macroscopic bone distractors. This system - the Microdistractor (MD) system - will be used to apply linear forces to ASCs derived from pediatric adipose tissue. In Specific Aim 1, the MD system will be optimized so that the osteogenic capacity of pediatric ASCs is maximized. In Specific Aim 2, these conditions will be used to pre-condition the ASCs followed by their implantation into cranial bony defects. Three percent of all newborns have significant bony abnormalities, including cleft palate and cranial vault defects. Current therapies for these defects are limited in their degree of healing. Moreover, the use of conventional bone-inducing growth factors like BMP2, may not be warranted in these cases. Therefore, this application describes the use of a novel distraction system that may promote the osteogenic capacity of ASCs in vitro without the need for exogenous growth factors. Public Health Relevance: Three percent of all infants born in the US each year exhibit some type of craniofacial defect, including the most commone defect - cleft palate. Tissue engineering strategies using the patient's own stem cells may allow surgeonsto correct such defects by augmenting the ability of these stem cells to undergo bone formation within the defect. One strategy to augment these stem cells may be the application of mechanical stresses to the cells in vitro prior to implantation within the defect, thus increasing their ability to heal substantial craniofacial defects.